Stroke is the fourth leading cause of death in the United States and lacks effective therapeutic interventions. The only FDA-approved treatment for acute ischemic stroke is intravenous tissue plasminogen activator (tPA), which rely heavily on the identification of the ischemic penumbra. The conventional criteria using MRI perfusion and diffusion mismatch as patient selection criterion for thrombolysis have not shown satisfactory results in several randomized trials. Magnetic Resonance Imaging (MRI) and Spectroscopy (MRS) hold great promise for the assessment of brain perfusion and metabolic status non-invasively and rapidly. However, there are still limited MRI techniques that can detect the metabolic change with high spatial resolution. Recently, so-called amide proton transfer weighted (APTw) Chemical Exchange Saturation Transfer (CEST) MRI has shown potential in detecting the consequence of early metabolic changes by making use of the pH-sensitive exchangeable amide protons in tissue proteins. However, the sensitivity of the APT method as a function of pH value is limited due to its low exchange rate. With this proposal, we attempt to develop a new CEST method, based on an on-resonance saturation scheme, which will allow us to map the brain pH values with high sensitivity by detecting the combined CEST signal from all types of fast-exchanging protons in proteins and metabolites, and will provide a novel MRI imaging module that can quantitatively assess tissue status for further treatment. Our recent publication had shown that the new CEST technique, on-resonance variable delay multiple pulse (onVDMP), is able to detect the fast-exchanging protons with high sensitivity. We, therefore, hypothesize that the CEST signal detected by onVDMP is associated with the pH values due to the abundant fast-exchanging protons in the protein and metabolic molecules. The optimized onVDMP 3D CEST method will be applied in mice in a transient stroke model to verify that the technique is sensitive enough to detect the changes in the metabolite level in the ischemic brain. At the same time, the MRI perfusion map and several histologic analyses will be performed to assess the anatomical brain injury. Upon the successful completion of this proposal, we anticipate developing a new clinic-ready MRI technique that detect and evaluate the metabolite change in the ischemic brain. !